ABSTRACT
Extracellular signal-regulated kinases (ERK) 1, 2 and 3 are involved in cell proliferation and differentiation, and apoptosis; although ERK1/2 have been widely studied, limited knowledge on ERK3 is available. The present work aimed at investigating ERK3 distribution during cell cycle and apoptosis in human tumor HeLa cells. The analysis performed by double immunofluorescence and immunoelectron microscopy experiments revealed that during interphase ERK3 is mainly resident in the nucleoplasm in association with ribonuclear proteins involved in early pre-mRNA splicing, it undergoes cell cycle-dependent redistribution and, during apoptosis, it remains in the nucleus in the form of massive nuclear aggregates, then moves to the cytoplasm and is finally extruded.
Subject(s)
Apoptosis/physiology , Cell Nucleus/enzymology , Cytoplasm/enzymology , Interphase/physiology , Mitogen-Activated Protein Kinase 6/metabolism , Mitosis/physiology , HeLa Cells , Humans , Protein Transport/physiologyABSTRACT
HMA (5-(N,N-hexamethylene)amiloride), which belongs to a family of novel amiloride derivatives, is one of the most effective inhibitors of Na+/H+ exchangers, while uneffective against Na+ channels and Na+/Ca2+ exchangers. In this study, we provided evidence that HMA can act as a fluorescent probe. In fact, human retinal ARPE19 cells incubated with HMA show an intense bluish fluorescence in the cytoplasm when observed at microscope under conventional UV-excitation conditions. Interestingly, a prolonged observation under continuous exposure to excitation lightdoes not induce great changes in cells incubated with HMA for times up to about 5 min, while an unexpected rapid increase in fluorescence signal is observed in cells incubated for longer times. The latter phenomenon is particularly evident in the perinuclear region and in discrete spots in the cytoplasm. Since HMA modulates intracellular acidity, the dependence of its fluorescence properties on medium pH and response upon irradiation have been investigated in solution, at pH 5.0 and pH 7.2. The changes in both spectral shape and amplitude emission indicate a marked pH influence on HMA fluorescence properties, making HMA exploitable as a self biomarker of pH alterations in cell studies, in the absence of perturbations induced by the administration of other exogenous dyes.
Subject(s)
Amiloride/analogs & derivatives , Fluorescent Dyes/chemistry , Sodium-Hydrogen Exchangers/antagonists & inhibitors , Amiloride/chemistry , Amiloride/pharmacology , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Cell Line , Fluorescent Dyes/pharmacology , Humans , Hydrogen-Ion Concentration , Microscopy, Fluorescence/methods , Ultraviolet RaysABSTRACT
The purpose of this paper is to quantify by in vitro experiment free radical activity in jaw bone demineralization processes. Sample were take and graphic interpolation of calcium ions undertake. The significance of the results is compared with the paper that are published in literature on the same subject.
